Metagenomic expression

The rapidly increasing number of novel metagenome-derived genes also led to the identiflcation of major bottlenecks. These are mainly linked to the eflhciency of heterologous gene expression and the amount of protein produced, in particular. 

In the metagenomic approach, DNA was directly extracted from uncultured samples followed by cloning and expression [3]. For example, by combination of directed evolution with the metagenome approach, an a-amylase mutant with optimal activity at pH 4.5 and optimal thermostability at 105 °C was discovered for starch liquefaction and EtOH production [4]. 

Figure 2.3 Metagenomic cloning experiments. Isolation of genomic DNA directly from environments (soil, plants, mixed environments or thermal-vent worms are the examples Illustrated here) can recover DNA fragments which could encode for enzymes. The DNA fragments can be ligated to plasmids or DNA linkers, and then subjected to functional screening (expression cloning) and/or sequence analysis. Amplification by PCR can sometimes be used to yield libraries enriched with clones containing selected sequence motifs relating to families of enzymes...

Cottrell, M. T., Yu, L. Y., and Kirchman, D. L. (2005). Sequence and expression analyses of Cytophaga-like hydrolases in a Western arctic metagenomic library and the Sargasso seat. Appl. Environ. Microbiol 71, 8506—8513. 

A number of metagenomic libraries hosted in E. coli have now been screened for clones that produce either antibacterial activities or color. In addition to hits that are unique to each library, these studies have encountered four common hits. These include the antibacterially active long-chain A-acyl amino acid-producing clones described earlier in this section, red antibacterially active clones that express aminolevulinic acid synthases (hemA), brown clones that produce melanin-like polymers, and blue clones that produce mixtures of indigo (25) and indinibin... 

Figure 10 Clones that produce either indigo (25) or indirubin (26) have been identified in three different metagenomic studies. The key features of these three studies and the molecules characterized in each study are outlined here. Two of the indigo-producing clones were identified in screens for color and antibacterial activity while the third was found in an assay for quorum-sensing mimics (METREX). In this intracellular assay for quorum-sensing inducers, activated LuxR (LuxRbound toan AHL or an AHL mimic) induces the expression of GFP. ...

Functionally expressed, 1626 Functional metagenomic, 2455 Functional properties, 3981 Functional redundancy, 2441 Functional value, 2855, 2862, 2871, 2874 Fungal, 1688, 1690, 1694, 1695, 1699 cell membrane, 3981 disease, 3976 elicitors, 2781,4045 pathogens, 2937... 

Metatranscriptomics, metaproteomics, and metabolomics are also widely used in conjunction with metagenomic analyses. These omic technologies can provide a more comprehensive understanding of the types of genes being expressed... 

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